Li Min, Zhou Jing, Jin Weifeng, Li Xiaohong, Zhang Yuyan
School of Life Science, Zhejiang Chinese Medical University, Hangzhou, China.
Front Pharmacol. 2018 Apr 6;9:308. doi: 10.3389/fphar.2018.00308. eCollection 2018.
Hemorrhagic transformation, neurotoxicity, short treatment time windows, and other defects are considered as the major limitations for the thrombolytic therapy. This study is devoted to figure out whether Danhong injection (DHI) combined with tissue-plasminogen activator (t-PA) could extend the treatment time windows and ameliorate brain injury, hemorrhagic complication and BBB disruption after focal embolic stroke. , the combined concentrations of DHI and t-PA were added to wells reacted with plasminogen and D-Val-Leu-Lys-AMC. The optimum ratio of the combination of DHI plus t-PA was explored by detecting relative fluorescent. experiments, we firstly investigated the optimal dose of t-PA and Danhong injection for focal embolic stroke. The neurological deficit score, infarct volume and brain edema were assessed. Secondly, we proved that the combination group extended the thrombolytic window for treatment of focal embolic stroke. The neurological deficit score, infarct volume, brain edema and hemorrhagic complication were assessed, while levels of BAX, Bcl-2 and caspase-3 in brain tissue were analyzed by real-time polymerase chain reaction. Finally, to ask whether combination therapy with DHI plus t-PA protected the blood-brain barrier in a rat model of focal embolic stroke, neurological deficit score, ELISA, RT-PCR, western blot and fluorescence were used to detect the indicators of blood-brain barrier, such as tight junction protein, blood-brain barrier permeability and related gene expression. , plasmin activity assays showed that the combination of t-PA with DHI at about 1:1.6 w/v ratio increased by almost 1.4-fold the plasmin-generating capability of t-PA. experiments, the results showed that the combination of Danhong injection (4 mL/kg) and t-PA (2.5 mg/kg) could extend the t-PA treatment time windows to 4.5 h. And the combination t-PA (2.5 mg/kg) with DHI (4 mL/kg) ameliorated neurological score, cerebral infarction, brain edema, brain hemorrhage, and BBB disruption. Combination therapy with Danhong injection (4 mL/kg) plus t-PA (2.5 mg/kg) could extend the t-PA treatment time windows to 4.5 h, ameliorate BBB disruption, reduce infarction, brain swelling and hemorrhage after ischemic stroke.
出血转化、神经毒性、治疗时间窗短等缺陷被认为是溶栓治疗的主要局限性。本研究旨在探究丹红注射液（DHI）联合组织型纤溶酶原激活剂（t-PA）是否能延长治疗时间窗，并改善局灶性栓塞性中风后的脑损伤、出血并发症和血脑屏障破坏。将DHI和t-PA的联合浓度加入与纤溶酶原和D-缬氨酸-亮氨酸-赖氨酸-7-氨基-4-甲基香豆素反应的孔中。通过检测相对荧光来探索DHI加t-PA联合的最佳比例。在实验中，我们首先研究了t-PA和丹红注射液对局灶性栓塞性中风的最佳剂量。评估神经功能缺损评分、梗死体积和脑水肿。其次，我们证明联合组延长了局灶性栓塞性中风治疗的溶栓时间窗。评估神经功能缺损评分、梗死体积、脑水肿和出血并发症，同时通过实时聚合酶链反应分析脑组织中BAX、Bcl-2和半胱天冬酶-3的水平。最后，为了探究DHI加t-PA联合治疗是否能在局灶性栓塞性中风大鼠模型中保护血脑屏障，使用神经功能缺损评分、酶联免疫吸附测定、逆转录-聚合酶链反应、蛋白质免疫印迹和荧光检测血脑屏障的指标，如紧密连接蛋白、血脑屏障通透性和相关基因表达。纤溶酶活性测定表明，t-PA与DHI以约1:1.6 w/v的比例联合可使t-PA产生纤溶酶的能力提高近1.4倍。在实验中，结果表明丹红注射液（4 mL/kg）和t-PA（2.5 mg/kg）联合可将t-PA治疗时间窗延长至4.5小时。并且t-PA（2.5 mg/kg）与DHI（4 mL/kg）联合改善了神经评分、脑梗死、脑水肿、脑出血和血脑屏障破坏。丹红注射液（4 mL/kg）加t-PA（2.5 mg/kg）联合治疗可将t-PA治疗时间窗延长至4.5小时，改善血脑屏障破坏，减少缺血性中风后的梗死、脑肿胀和出血。
